As SARS-CoV-2 infections and death counts continue to rise, it remains unclear why some individuals recover from infection, whereas others rapidly progress and die. Although the immunological ...mechanisms that underlie different clinical trajectories remain poorly defined, pathogen-specific antibodies often point to immunological mechanisms of protection. Here, we profiled SARS-CoV-2-specific humoral responses in a cohort of 22 hospitalized individuals. Despite inter-individual heterogeneity, distinct antibody signatures resolved individuals with different outcomes. Although no differences in SARS-CoV-2-specific IgG levels were observed, spike-specific humoral responses were enriched among convalescent individuals, whereas functional antibody responses to the nucleocapsid were elevated in deceased individuals. Furthermore, this enriched immunodominant spike-specific antibody profile in convalescents was confirmed in a larger validation cohort. These results demonstrate that early antigen-specific and qualitative features of SARS-CoV-2-specific antibodies point to differences in disease trajectory, highlighting the potential importance of functional antigen-specific humoral immunity to guide patient care and vaccine development.
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•Limited early differences were observed in titers and neutralization across groups•Five antibody features could collectively differentiate convalescents and deceased•A shift in the balance of spike versus nucleocapsid immunity separated the groups•Spike-specific phagocytic and complement fixing activity was enriched in convalescents
Although most SARS-CoV-2-infected individuals experience mild disease, a significant fraction of individuals become severely infected. Early biomarkers that predict outcome are urgently needed. Atyeo et al. demonstrate that distinct acute SARS-CoV-2 humoral immune responses exist across severely ill individuals who ultimately convalesce or pass away.
Vaccines for rapidly evolving pathogens will confer lasting immunity if they elicit antibodies recognizing conserved epitopes, such as a receptor-binding site (RBS). From characteristics of an ...influenza-virus RBS-directed antibody, we devised a signature motif to search for similar antibodies. We identified, from three vaccinees, over 100 candidates encoded by 11 different VH genes. Crystal structures show that antibodies in this class engage the hemagglutinin RBS and mimic binding of the receptor, sialic acid, by supplying a critical dipeptide on their projecting, heavy-chain third complementarity determining region. They share contacts with conserved, receptor-binding residues but contact different residues on the RBS periphery, limiting the likelihood of viral escape when several such antibodies are present. These data show that related modes of RBS recognition can arise from different germline origins and mature through diverse affinity maturation pathways. Immunogens focused on an RBS-directed response will thus have a broad range of B cell targets.
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•Identified a signature motif for influenza RBS-directed antibodies•Influenza RBS-antibodies are nearly unrestricted in gene usage•Multiple donors have RBS-directed antibodies with receptor mimicry•Viral resistance to one RBS antibody does not confer resistance to all
A structure-based signature motif found in many human antibodies indicates recogition of the receptor-binding site of influenza virus hemagglutinin. These antibodies arise from diverse germline origins and affinity maturation pathways and suggest potential routes to a universal flu vaccine.
The diagnosis of COVID‐19 requires integration of clinical and laboratory data. Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) diagnostic assays play a central role in diagnosis and ...have fixed technical performance metrics. Interpretation becomes challenging because the clinical sensitivity changes as the virus clears and the immune response emerges. Our goal was to examine the clinical sensitivity of two most common SARS‐CoV‐2 diagnostic test modalities, polymerase chain reaction (PCR) and serology, over the disease course to provide insight into their clinical interpretation in patients presenting to the hospital. We conducted a single‐center, retrospective study. To derive clinical sensitivity of PCR, we identified 209 PCR‐positive SARS‐CoV‐2 patients with multiple PCR test results (624 total PCR tests) and calculated daily sensitivity from date of symptom onset or first positive test. Clinical sensitivity of PCR decreased with days post symptom onset with >90% clinical sensitivity during the first 5 days after symptom onset, 70%‐71% from Days 9 to 11, and 30% at Day 21. To calculate daily clinical sensitivity by serology, we utilized 157 PCR‐positive patients with a total of 197 specimens tested by enzyme‐linked immunosorbent assay for IgM, IgG, and IgA anti‐SARS‐CoV‐2 antibodies. In contrast to PCR, serological sensitivity increased with days post symptom onset with >50% of patients seropositive by at least one antibody isotype after Day 7, >80% after Day 12, and 100% by Day 21. Taken together, PCR and serology are complimentary modalities that require time‐dependent interpretation. Superimposition of sensitivities over time indicate that serology can function as a reliable diagnostic aid indicating recent or prior infection.
Germinal center (GC) B cells evolve toward increased affinity by a Darwinian process that has been studied primarily in genetically restricted, hapten-specific responses. We explored the population ...dynamics of genetically diverse GC responses to two complex antigens—Bacillus anthracis protective antigen and influenza hemagglutinin—in which B cells competed both intra- and interclonally for distinct epitopes. Preferred VH rearrangements among antigen-binding, naive B cells were similarly abundant in early GCs but, unlike responses to haptens, clonal diversity increased in GC B cells as early “winners” were replaced by rarer, high-affinity clones. Despite affinity maturation, inter- and intraclonal avidities varied greatly, and half of GC B cells did not bind the immunogen but nonetheless exhibited biased VH use, V(D)J mutation, and clonal expansion comparable to antigen-binding cells. GC reactions to complex antigens permit a range of specificities and affinities, with potential advantages for broad protection.
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•Developed a single B cell culture to characterize the mouse B cell repertoire•Affinity maturation of GC B cells is accompanied by increased VH representation•GCs permit a wide range of intra- and interclonal BCR affinities•Substantial fractions of GC B cells do not detectably bind native antigen
B cell selection and affinity maturation in germinal centers (GCs) elicited by complex antigens are not well studied. Kelsoe and colleagues demonstrate that affinity maturation in GCs is accompanied by increased clonal diversity and large variances in inter- and intraclonal BCR avidities in response to anthrax protective antigen and influenza hemagglutinin.
As the COVID-19 pandemic continues to spread, investigating the processes underlying the interactions between SARS-CoV-2 and its hosts is of high importance. Here, we report the identification of ...CD209L/L-SIGN and the related protein CD209/DC-SIGN as receptors capable of mediating SARS-CoV-2 entry into human cells. Immunofluorescence staining of human tissues revealed prominent expression of CD209L in the lung and kidney epithelia and endothelia. Multiple biochemical assays using a purified recombinant SARS-CoV-2 spike receptor-binding domain (S-RBD) or S1 encompassing both N termal domain and RBD and ectopically expressed CD209L and CD209 revealed that CD209L and CD209 interact with S-RBD. CD209L contains two N-glycosylation sequons, at sites N92 and N361, but we determined that only site N92 is occupied. Removal of the N-glycosylation at this site enhances the binding of S-RBD with CD209L. CD209L also interacts with ACE2, suggesting a role for heterodimerization of CD209L and ACE2 in SARS-CoV-2 entry and infection in cell types where both are present. Furthermore, we demonstrate that human endothelial cells are permissive to SARS-CoV-2 infection, and interference with CD209L activity by a knockdown strategy or with soluble CD209L inhibits virus entry. Our observations demonstrate that CD209L and CD209 serve as alternative receptors for SARS-CoV-2 in disease-relevant cell types, including the vascular system. This property is particularly important in tissues where ACE2 has low expression or is absent and may have implications for antiviral drug development.
The predominant approach for antibody generation remains animal immunization, which can yield exceptionally selective and potent antibody clones owing to the powerful evolutionary process of somatic ...hypermutation. However, animal immunization is inherently slow, not always accessible and poorly compatible with many antigens. Here, we describe ‘autonomous hypermutation yeast surface display’ (AHEAD), a synthetic recombinant antibody generation technology that imitates somatic hypermutation inside engineered yeast. By encoding antibody fragments on an error-prone orthogonal DNA replication system, surface-displayed antibody repertoires continuously mutate through simple cycles of yeast culturing and enrichment for antigen binding to produce high-affinity clones in as little as two weeks. We applied AHEAD to generate potent nanobodies against the SARS-CoV-2 S glycoprotein, a G-protein-coupled receptor and other targets, offering a template for streamlined antibody generation at large.Autonomous hypermutation yeast surface display (AHEAD) mimics the process of somatic hypermutation in animals to enable the rapid in vitro evolution of antibodies, including nanobodies targeting the RBD of SARS-CoV-2.
After vaccination or infection, long-lived germinal centers can produce antibodies with high affinity and specificity against pathogens. In this issue of Cell, de Carvalho et al. and Hägglöf et al. ...show that naive B cells can invade germinal centers, replacing B cells that entered early and changing features of antibody production. These findings have implications for vaccine design.
After vaccination or infection, long-lived germinal centers can produce antibodies with high affinity and specificity against pathogens. In this issue of Cell, de Carvalho et al. and Hägglöf et al. show that naive B cells can invade germinal centers, replacing B cells that entered early and changing features of antibody production. These findings have implications for vaccine design.
Vaccines to generate durable humoral immunity against antigenically evolving pathogens such as the influenza virus must elicit antibodies that recognize conserved epitopes. Analysis of single memory ...B cells from immunized human donors has led us to characterize a previously unrecognized epitope of influenza hemagglutinin (HA) that is immunogenic in humans and conserved among influenza subtypes. Structures show that an unrelated antibody from a participant in an experimental infection protocol recognized the epitope as well. IgGs specific for this antigenic determinant do not block viral infection in vitro, but passive administration to mice affords robust IgG subtype-dependent protection against influenza infection. The epitope, occluded in the pre-fusion form of HA, is at the contact surface between HA head domains; reversible molecular “breathing” of the HA trimer can expose the interface to antibody and B cells. Antigens that present this broadly immunogenic HA epitope may be good candidates for inclusion in “universal” flu vaccines.
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•Human B cells specific for a novel epitope on influenza A groups 1 and 2•Crystallography locates the epitope at the interface of the hemagglutinin head domains•Robust protection by antibodies to this epitope, dependent on IgG subclass•Protective, cross-group antibodies are encoded by diverse sets of Ig gene segments
Antibodies targeting a novel site at the interface of the head domains of hemagglutinin provide broad, IgG-subclass-dependent protection against influenza.
The SARS-CoV-2 pandemic has affected more than 185 million people worldwide resulting in over 4 million deaths. To contain the pandemic, there is a continued need for safe vaccines that provide ...durable protection at low and scalable doses and can be deployed easily. Here, AAVCOVID-1, an adeno-associated viral (AAV), spike-gene-based vaccine candidate demonstrates potent immunogenicity in mouse and non-human primates following a single injection and confers complete protection from SARS-CoV-2 challenge in macaques. Peak neutralizing antibody titers are sustained at 1 year and complemented by functional memory T cell responses. The AAVCOVID vector has no relevant pre-existing immunity in humans and does not elicit cross-reactivity to common AAVs used in gene therapy. Vector genome persistence and expression wanes following injection. The single low-dose requirement, high-yield manufacturability, and 1-month stability for storage at room temperature may make this technology well suited to support effective immunization campaigns for emerging pathogens on a global scale.
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•AAVCOVID is a gene- and AAV-based spike antigen COVID-19 vaccine•A single AAVCOVID dose provides protection from SARS-CoV-2 challenge in monkeys•Neutralizing antibody levels in animals are maintained at peak for a year•AAVCOVID can be produced at scale and stored at room temperature for 1 month
Zabaleta et al. report AAVCOVID, an experimental preventative vaccine for COVID-19. AAVCOVID is a vaccine that delivers the SARS-CoV-2 spike antigen as a gene via an adeno-associated vector. Results demonstrate protection from viral challenge in NHPs, durable immunogenicity, cross-reactivity with emerging variants, a scalable production method, and room-temperature stability for 1 month.
Epitope-focused approaches for selective clonal induction of broadly neutralizing antibodies (bnAbs) inform most current vaccine strategies for influenza virus and other rapidly evolving pathogens. ...The two conserved epitopes on the influenza hemagglutinin (HA) - the "stem" and the receptor-binding site (RBS) on the "head" - are the focus of the current "universal" influenza vaccine development efforts. Because stem-directed serum bnAbs are much less abundant than head-directed ones, we hypothesized that the HA stem bnAbs may be autoreactive and thus eliminated through the mechanisms of self-tolerance. We compared autoreactivity profiles of a set of stem and head-directed bnAbs. Most of the stem bnAbs we examined bound autoantigens; several showed staining of HEp-2 cells. A smaller proportion of the head-directed bnAbs were polyreactive. Gene usage did not correlate with autoreactivity. We suggest that complex foreign antigens may often have surface patches resembling some host epitope; our results indicate that HA stem epitopes resemble a host epitope more frequently than does the RBS.
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